| 1. |
- Alm, Kersti, et al.
(författare)
-
Digital holography and cell studies
- 2011
-
Ingår i: Holography, Research and Technologies. - : DKV - Deutscher Kälte- und Klimatechnischer Verein. - 9789533072272 ; , s. 237-252
-
Bokkapitel (övrigt vetenskapligt/konstnärligt)abstract
- Digital holography microscopy (DHM) has developed into a broad field, and one of all the interesting applications is to study cells without staining, labeling or in any other way affecting them. Both fixed and living, dying or dead cells can be studied. The first DHM images showing living cells were published in 2004 and 2005 (Carl et al. 2004, Marquet et al. 2005), making this field of research rather new. Digital holography makes it possible to easily measure cell properties that previously have been very difficult to study, such as cell thickness and volume (Marquet et al. 2005, Mölder et al. 2008). Two of the major advantages of DHM is the 3-D imaging possibility and measurements over time. Digital holography has ben used to study several types of cells, such as nerve cells, red blood cells, stem cells and cancer cells (Emery et al. 2007, Kemper et al. 2006, Langehanenberg et al. 2009) . It has also been applied for studies of cell proliferation, cell movement, sub-cellular structures and cell morphology (Kemper et al. 2009, Yu et al. 2009). Both 2-D and 3-D cell movement can be determined ( Langehanenberg et al. 2009). Even cell viability status can be determined using DHM. Interestingly, it is possible to study both single cells and entire populations simultaneously, allowing for very nuanced studies. Older, well known techniques often require some degree of cell disturbance such as the fluorescent antibody labeling required for fluorescense or confocal microscopy studies. In this paper we will present some of the studies made possible by DHM. We will compare DHM with previously used techniques and discuss the benefits and drawbacks of digital holography cell measurements.
|
|
| 2. |
- Cirenajwis, Helena, et al.
(författare)
-
Molecular stratification of metastatic melanoma using gene expression profiling: prediction of survival outcome and benefit from molecular targeted therapy.
- 2015
-
Ingår i: Oncotarget. - : Impact Journals, LLC. - 1949-2553. ; 6:14, s. 12297-12309
-
Tidskriftsartikel (refereegranskat)abstract
- Melanoma is currently divided on a genetic level according to mutational status. However, this classification does not optimally predict prognosis. In prior studies, we have defined gene expression phenotypes (high-immune, pigmentation, proliferative and normal-like), which are predictive of survival outcome as well as informative of biology. Herein, we employed a population-based metastatic melanoma cohort and external cohorts to determine the prognostic and predictive significance of the gene expression phenotypes. We performed expression profiling on 214 cutaneous melanoma tumors and found an increased risk of developing distant metastases in the pigmentation (HR, 1.9; 95% CI, 1.05-3.28; P=0.03) and proliferative (HR, 2.8; 95% CI, 1.43-5.57; P=0.003) groups as compared to the high-immune response group. Further genetic characterization of melanomas using targeted deep-sequencing revealed similar mutational patterns across these phenotypes. We also used publicly available expression profiling data from melanoma patients treated with targeted or vaccine therapy in order to determine if our signatures predicted therapeutic response. In patients receiving targeted therapy, melanomas resistant to targeted therapy were enriched in the MITF-low proliferative subtype as compared to pre-treatment biopsies (P=0.02). In summary, the melanoma gene expression phenotypes are highly predictive of survival outcome and can further help to discriminate patients responding to targeted therapy.
|
|
| 3. |
- Cirenajwis, Helena
(författare)
-
Molecular Subtypes of Melanoma. Biological and Clinical Significance.
- 2016
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Cutaneous malignant melanoma (CMM) is the most lethal form of skin cancer and its incidence has increased faster than that of any other cancer, rendering it a major public health problem worldwide. High-throughput screenings have opened the door to a new scientific world, which enables molecular-based characterization of large cancer cohort collections. The aim of the research presented in this thesis was to explore the molecular landscapes of melanoma tumors on a genomic and transcriptomic level and subsequently correlate certain molecular features with patient survival, treatment response and tumor evolutionary patterns. In Paper I, it was concluded that metastatic melanoma could be divided into transcriptomic subtypes (gene expression (GEX) phenotypes) possessing diverse biological and clinical features. Patients harboring melanomas infiltrated by immune cells, i.e. the high-immune subtype, showed a superior survival, whereas highly proliferative melanomas, i.e. the proliferative subtype, was correlated to a poor survival outcome and resistance to targeted therapies. Moreover, it was also shown that, irrespectively of the GEX phenotypes, melanomas could be divided into genomic subtypes based on genetic aberrations in the mitogen-activated protein kinase (MAPK) signaling pathway. In Paper II, it was found that mutations in the tumor suppressor gene neurofibromin 1 (NF1), was linked to inferior survival. Today, it is well accepted that most tumors possess some level of intratumor heterogeneity (ITH), i.e. subclonality, influencing disease progression. In Papers III and IV, the evolutionary aspects of melanoma were considered by analyzing ITH, as well as disease progression on a molecular basis. When analyzing multiple metastatic lesions from individual patients, we found that most tumors were genetically different, with a common stem of genetic aberrations and the addition of new “private” ones, thus pointing to continued evolution during progression. Moreover, the GEX proliferative phenotype appeared to be correlated to a later disease course. From multiregional biopsies from single tumors, it was found that mutations in the MAPK signaling pathway appeared to be early events in tumorigenesis. Heterogeneous somatic mutations were found in the range of 3-38%, thus highlighting different levels of subclonality in melanoma. A high degree of mutational heterogeneity was associated with a more aggressive disease progression. In conclusion, melanoma is a complex molecular disease that can be characterized by genomic and transcriptomic signatures with clinical implications. However, a single biopsy might not reflect the true tumor complexity, and subclonality may be one reason behind resistance development.
|
|
| 4. |
- Cirenajwis, Helena, et al.
(författare)
-
NF1-mutated melanoma tumors harbor distinct clinical and biological characteristics
- 2017
-
Ingår i: Molecular Oncology. - : Wiley. - 1574-7891. ; 11:4, s. 438-451
-
Tidskriftsartikel (refereegranskat)abstract
- In general, melanoma can be considered as a UV-driven disease with an aggressive metastatic course and high mutational load, with only few tumors (acral, mucosal, and uveal melanomas) not induced by sunlight and possessing a lower mutational load. The most commonly activated pathway in melanoma is the mitogen-activated protein kinase (MAPK) pathway. However, the prognostic significance of mutational stratification is unclear and needs further investigation. Here, in silico we combined mutation data from 162 melanomas subjected to targeted deep sequencing with mutation data from three published studies. Tumors from 870 patients were grouped according to BRAF, RAS, NF1 mutation or triple-wild-type status and correlated with tumor and patient characteristics. We found that the NF1-mutated subtype had a higher mutational burden and strongest UV mutation signature. Searching for co-occurring mutated genes revealed the RASopathy genes PTPN11 and RASA2, as well as another RAS domain-containing gene RASSF2 enriched in the NF1 subtype after adjustment for mutational burden. We found that a larger proportion of the NF1-mutant tumors were from males and with older age at diagnosis. Importantly, we found an increased risk of death from melanoma (disease-specific survival, DSS; HR, 1.9; 95% CI, 1.21-3.10; P = 0.046) and poor overall survival (OS; HR, 2.0; 95% CI, 1.28-2.98; P = 0.01) in the NF1 subtype, which remained significant after adjustment for age, gender, and lesion type (DSS P = 0.03, OS P = 0.06, respectively). Melanoma genomic subtypes display different biological and clinical characteristics. The poor outcome observed in the NF1 subtype highlights the need for improved characterization of this group.
|
|
| 5. |
- Cirenajwis, Helena, et al.
(författare)
-
Performance of gene expression-based single sample predictors for assessment of clinicopathological subgroups and molecular subtypes in cancers : a case comparison study in non-small cell lung cancer
- 2019
-
Ingår i: Briefings in Bioinformatics. - : Oxford University Press (OUP). - 1477-4054 .- 1467-5463. ; 21:2, s. 729-740
-
Tidskriftsartikel (refereegranskat)abstract
- The development of multigene classifiers for cancer prognosis, treatment prediction, molecular subtypes or clinicopathological groups has been a cornerstone in transcriptomic analyses of human malignancies for nearly two decades. However, many reported classifiers are critically limited by different preprocessing needs like normalization and data centering. In response, a new breed of classifiers, single sample predictors (SSPs), has emerged. SSPs classify samples in an N-of-1 fashion, relying on, e.g. gene rules comparing expression values within a sample. To date, several methods have been reported, but there is a lack of head-to-head performance comparison for typical cancer classification problems, representing an unmet methodological need in cancer bioinformatics. To resolve this need, we performed an evaluation of two SSPs [k-top-scoring pair classifier (kTSP) and absolute intrinsic molecular subtyping (AIMS)] for two case examples of different magnitude of difficulty in non-small cell lung cancer: gene expression–based classification of (i) tumor histology and (ii) molecular subtype. Through the analysis of ~2000 lung cancer samples for each case example (n = 1918 and n = 2106, respectively), we compared the performance of the methods for different sample compositions, training data set sizes, gene expression platforms and gene rule selections. Three main conclusions are drawn from the comparisons: both methods are platform independent, they select largely overlapping gene rules associated with actual underlying tumor biology and, for large training data sets, they behave interchangeably performance-wise. While SSPs like AIMS and kTSP offer new possibilities to move gene expression signatures/predictors closer to a clinical context, they are still importantly limited by the difficultness of the classification problem at hand.
|
|
| 6. |
- Cirenajwis, Helena, et al.
(författare)
-
Reduction of the putative CD44+CD24- breast cancer stem cell population by targeting the polyamine metabolic pathway with PG11047.
- 2010
-
Ingår i: Anti-Cancer Drugs. - 0959-4973. ; 21:10, s. 897-906
-
Tidskriftsartikel (refereegranskat)abstract
- Cancer stem cells (CSCs) are considered to be of particular concern in cancer as they possess inherent properties of self-renewal and differentiation, along with expressing certain genes related to a mesenchymal phenotype. These features favour the promotion of tumour recurrence and metastasis in cancer patients. Thus, the optimal chemotherapeutic treatment should target the CSC population, either by killing these cells and/or by inducing their transition to a more differentiated epithelial-like phenotype. Experiments were carried out on the trastuzumab-resistant human epidermal growth factor receptor 2-overexpressing breast cancer cell line JIMT-1 to unravel the chemotherapeutic effects of the polyamine analogue [N,N]bis(ethyl)-cis-6,7-dehydrospermine (PG11047) and of the polyamine biosynthetic inhibitor 2-difluoromethylornithine (DFMO) on the CD44CD24 CSC population. Furthermore, effects on the properties of self-renewal and epithelial/mesenchymal markers were also investigated. Treatment with PG11047 reduced the CD44CD24 subpopulation of JIMT-1 cells by approximately 50%, inhibited and/or reduced self-renewal capability of the CSC population, decreased cell motility and induced expression of mesenchymal to epithelial transition-associated proteins that are involved in promoting an epithelial phenotype. By contrast, DFMO slightly increased the CD44CD24 subpopulation, increased cell motility and the level of mesenchymal-related proteins. DFMO treatment reduced the self-renewal capability of the CSC population. Both PG11047 and DFMO reduced the expression of the human epidermal growth factor receptor 2 protein, which is correlated to malignancy and resistance to trastuzumab in JIMT-1 cells. Our findings indicate that treatment with PG11047 targeted the CSC population by interfering with several stem cell-related properties, such as self-renewal, differentiation, motility and the mesenchymal phenotype.
|
|
| 7. |
|
|
| 8. |
- Harbst, Katja, et al.
(författare)
-
Molecular and genetic diversity in the metastatic process of melanoma.
- 2014
-
Ingår i: Journal of Pathology. - : Wiley. - 0022-3417 .- 1096-9896. ; 233:1, s. 39-50
-
Tidskriftsartikel (refereegranskat)abstract
- Diversity between metastatic melanoma tumours in individual patients is known; however, the molecular and genetic differences remain unclear. To examine the molecular and genetic differences between metastatic tumours, we performed gene-expression profiling of 63 melanoma tumours obtained from 28 patients (two or three tumours/patient), followed by analysis of their mutational landscape, using targeted deep sequencing of 1697 cancer genes and DNA copy number analysis. Gene-expression signatures revealed discordant phenotypes between tumour lesions within a patient in 50% of the cases. In 18 of 22 patients (where matched normal tissue was available), we found that the multiple lesions within a patient were genetically divergent, with one or more melanoma tumours harbouring 'private' somatic mutations. In one case, the distant subcutaneous metastasis of one patient occurring 3 months after an earlier regional lymph node metastasis had acquired 37 new coding sequence mutations, including mutations in PTEN and CDH1. However, BRAF and NRAS mutations, when present in the first metastasis, were always preserved in subsequent metastases. The patterns of nucleotide substitutions found in this study indicate an influence of UV radiation but possibly also DNA alkylating agents. Our results clearly demonstrate that metastatic melanoma is a molecularly highly heterogeneous disease that continues to progress throughout its clinical course. The private aberrations observed on a background of shared aberrations within a patient provide evidence of continued evolution of individual tumours following divergence from a common parental clone, and might have implications for personalized medicine strategies in melanoma treatment. Published by John Wiley & Sons, Ltd. www.pathsoc.org.uk.
|
|
| 9. |
- Harbst, Katja, et al.
(författare)
-
Multiregion whole-exome sequencing uncovers the genetic evolution and mutational heterogeneity of early-stage metastatic melanoma
- 2016
-
Ingår i: Cancer Research. - 0008-5472. ; 76:16, s. 4765-4774
-
Tidskriftsartikel (refereegranskat)abstract
- Cancer genome sequencing has shed light on the underlying genetic aberrations that drive tumorigenesis. However, current sequencing-based strategies, which focus on a single tumor biopsy, fail to take into account intratumoral heterogeneity. To address this challenge and elucidate the evolutionary history of melanoma, we performed whole-exome and transcriptome sequencing of 41 multiple melanoma biopsies from eight individual tumors. This approach revealed heterogeneous somatic mutations in the range of 3%-38% in individual tumors. Known mutations in melanoma drivers BRAF and NRAS were always ubiquitous events. Using RNA sequencing, we found that the majority of mutations were not expressed or were expressed at very low levels, and preferential expression of a particular mutated allele did not occur frequently. In addition, we found that the proportion of ultraviolet B (UVB) radiation-induced C>T transitions differed significantly (P <0.001) between early and late mutation acquisition, suggesting that different mutational processes operate during the evolution of metastatic melanoma. Finally, clinical history reports revealed that patients harboring a high degree of mutational heterogeneity were associated with more aggressive disease progression. In conclusion, our multiregion tumor-sequencing approach highlights the genetic evolution and non-UVB mutational signatures associated with melanoma development and progression, and may provide a more comprehensive perspective of patient outcome.
|
|
| 10. |
- Howlin, Jillian, et al.
(författare)
-
Loss of CITED1, an MITF regulator, drives a phenotype switch in vitro and can predict clinical outcome in primary melanoma tumours.
- 2015
-
Ingår i: PeerJ. - : PeerJ. - 2167-8359. ; 3
-
Tidskriftsartikel (refereegranskat)abstract
- CITED1 is a non-DNA binding transcriptional co-regulator whose expression can distinguish the 'proliferative' from 'invasive' signature in the phenotype-switching model of melanoma. We have found that, in addition to other 'proliferative' signature genes, CITED1 expression is repressed by TGFβ while the 'invasive' signature genes are upregulated. In agreement, CITED1 positively correlates with MITF expression and can discriminate the MITF-high/pigmentation tumour molecular subtype in a large cohort (120) of melanoma cell lines. Interestingly, CITED1 overexpression significantly suppressed MITF promoter activation, mRNA and protein expression levels while MITF was transiently upregulated following siRNA mediated CITED1 silencing. Conversely, MITF siRNA silencing resulted in CITED1 downregulation indicating a reciprocal relationship. Whole genome expression analysis identified a phenotype shift induced by CITED1 silencing and driven mainly by expression of MITF and a cohort of MITF target genes that were significantly altered. Concomitantly, we found changes in the cell-cycle profile that manifest as transient G1 accumulation, increased expression of CDKN1A and a reduction in cell viability. Additionally, we could predict survival outcome by classifying primary melanoma tumours using our in vitro derived 'CITED1-silenced' gene expression signature. We hypothesize that CITED1 acts a regulator of MITF, functioning to maintain MITF levels in a range compatible with tumourigenesis.
|
|
